The invention relates to a method for diagnosing certain pathological conditions, particularly vaginal disorders, by determining biogenic amines in samples of body fluids or other samples and distinguishing between more volatile and the less volatile amine species. The invention also includes selectively enhancing the emanation of the less volatile species and performing a partial separation between the more volatile and the less volatile species. The invention also relates to an apparatus for automatically providing diagnoses of certain pathological conditions and/or relevant diagnostic information.
It is known in the art that the presence of biogenic amines in human body fluids may reveal or suggest pathological conditions and dysfunctions. For example, elevated levels of certain biogenic amines in urine may indicate the presence or the likelihood of the presence of a cancer (there are many papers dealing with this—see, for instance, Suh, J W, Lee, S H, Chung, B C, Park, J, Urinary Polyamine Evaluation for Effective Diagnosis of Various Cancers, Journal of Chromatography B, 1997, Vol. 688, Iss 2, pp. 179-186). Several of the types of vaginal diseases may be expressed in elevated levels of biogenic amines in vaginal discharge and fluids (see, for instance, C. S. Chen, R. Amsel, D. A. Eschenbach and K. K. Holmes, Biochemical diagnosis of vaginitis: determination of diamines in vaginal fluid, J. Infectious Disease 145 (1982), pp. 337-345).
Body fluids may include e.g. urine, blood, serum, saliva, vaginal discharge and fluids, etc. Further, samples in which the presence of biogenic amines may be determined may not be fluids, but, e.g., skin and tissues, swipe samples, etc. Even direct sniffing of skin or breath exhaled by a subject may provide information in this respect. This should be understood whenever body fluids are mentioned in this application.
Chemical changes in the living system or degradation processes of cells after death are accompanied with formation of molecular byproducts. These processes include the breaking down of peptides and DNA strands to smaller components, and changes in the building blocks, amino acids, leading to the formation of amines. Not only amino compounds are produced, but other, smaller molecules, such as aldehydes and alcohols are also formed. One of the processes of particular interest is the breakdown of amino acids and the production of amines, diamines and polyamines. For example, decarboxylation of histidine, ornithine, lysine, produces histamine, putrescine and cadaverine, respectively.
Several analytical methods have been proposed for the analysis of biogenic amines. Most of these are laboratory methods that require expensive equipment, extensive sample preparation and the skills of a trained analytical chemist or technician. Among these are high performance liquid chromatography (HPLC), or gas chromatography after derivatization of the samples. Biosensors may also be used, as well as various spectrometric techniques. Solid state sensors have also been proposed, but generally lack specificity.
Ion Mobility Spectrometry (also, briefly, IMS) is a known analytical method and its application for the determination of aliphatic and aromatic amines has been suggested: see, for instance, Z. Karpas, Ion Mobility Spectrometry of Aliphatic and Aromatic Amines, Anal. Chem. 61 (1989), 684. An apparatus for carrying out this method—the Ion Mobility Spectrometer (IMS)—is used primarily for detection, identification and monitoring of trace amounts of gases and vapors. It is particularly suitable for detection of compounds that have high proton affinity and form stable positive ions, or for compounds that have a high electronegativity and readily form stable negative ions. IMS is fully discussed in J. I. Baumbach and G. A. Eiceman, Appl. Spectrosc. 1999, vol. 53, pp. 338A-355A. However, any device that may be used for determining or measuring the mobility of ions may be used for carrying out the invention, and therefore any reference to IMS in this description and claims should not be construed as a limitation, but should be construed any including instrument for determining or measuring the mobility of ions.
The knowledge of the prior art as to the importance of biogenic amines for the possible detection of pathological conditions and as to the analysis of biogenic amines, including the use of IMS, has failed so far to provide a simple and reliable method for the diagnosis of vaginal disorders, particularly, though not exclusively, bacterial vaginosis that affects a large number of women. The provision of such a diagnostic method would constitute a valuable contribution to the medical art. However, such a method is not available: the detection of biogenic amines in vaginal fluids is known to suggest the presence of a pathological condition, but it does not provide specific and reliable information and merely suggests to the specialized physician the desirability of carrying out whatever tests and examinations may finally lead to a diagnosis.
Samples collected from vegetation, clinical or biological media (the samples) contain biogenic amines that arise from degradation of amino acids through enzymatic and microbial processes. The types of biogenic amines and their quantities are indicative of the progress of said degradation processes, and can thus serve as a measure for food spoilage or the existence and extent of pathological conditions.
Some analytical methods for measuring biogenic amines, such as gas chromatography (GC) or ion mobility spectrometry (IMS), are based on determining their amount in the gas phase. However, while some of these biogenic amines are highly volatile, like trimethylamine (TMA), most have very low vapor pressures at room temperature, and are hardly present in the gas phase. Another reason for this is that biogenic amines may be present in the samples as salts, such as a hydrochloride form, and not present in the more volatile free-base structure. In such cases, their vapor pressure may be very low, and their detection and quantification by gas phase techniques may be very difficult.
One common practice to overcome such difficulties is to dissociate the salts and produce the free-bases by addition of an alkaline solution. For example, in carrying out the Amsel test for detection of bacterial vaginosis (a common vaginal infection), the physician adds a drop of 10% KOH solution to a swab collected from the vaginal fluid and tries to detect the presence of volatile amines by sniffing the sample (the so-called “whiff test”).
Further, the present knowledge does not provide the practicing physician with an apparatus for the quick diagnosis of bacterial vaginosis and other pathological conditions, by a simple and direct way and without the application of knowledge and technology that are typical of different branches of science and are not found together in any physician, no matter how competent and dedicated. It would be extremely valuable to provide an instrument and method whereby the average physician could obtain from bodily fluids, quickly and in a reliable way, a diagnostic indication of specific diseases and/or pathological conditions, even though such a diagnostic indication may not be final and conclusive and may require, whether positive or negative, verification and integration.
It is therefore a purpose of this invention to provide method for the diagnosis of vaginal disorders, particularly, though not exclusively, bacterial vaginosis.
It is another purpose to provide a method for carrying out such diagnosis automatically and in real time.
It is a further purpose to provide, automatically and in real time, information of fundamental value in the diagnosis of a variety of pathological conditions.
It is a still further purpose to overcome the problems arising in the detection of less volatile or semi-volatile amines, not only in clinical samples, but also in any vegetal or biological medium.
It is a still further purpose to selectively enhance the detection of less volatile or semi-volatile biogenic amines.
It is a still further purpose of this invention to permit to separate, at least partially, the more volatile from the less volatile amines.
It is a still further purpose to provide an apparatus for the quick diagnosis of bacterial vaginosis and other pathological conditions.
It is a still further purpose to provide a portable instrument that is capable of carrying out the spectrometry of bodily fluids and automatically derive from said spectrometry significant diagnostic indications.
It is a still further purpose to provide such an instrument that can be directed to provide diagnostic indications for specific diseases and/or pathological conditions.
It is a still further purpose to provide such an instrument that consists of the combination of components known in the art and readily available.
It is a still further purpose to provide such an instrument that can be widely used by physicians and medical institutions and is not excessively expensive.
It is a still further purpose to provide such an instrument which can be used, with the due precautions and warnings, by persons other than physicians and even by the patients themselves.
It is a still further purpose to provide such an instrument which can be used for purposes that are not diagnostic purposes, but are relevant to the public health, for instance, the control of the condition of food, such as, but not exclusively, the freshness of meat, fish and their products, as well as seafood.
It is a still further purpose to provide such an instrument that permits instant examination of tissues removed during an operation, or medical procedure, as an indication for malignant tissues.
Other purposes and advantages of the invention will appear as the description proceeds.